A single Cys706 to Phe substitution in the retinoblastoma protein causes the loss of binding to SV40 T antigen.
Most naturally occurring mutants of the retinoblastoma (RB) protein contain large deletions or truncations. The small cell lung carcinoma cell line H209 contains a normal-sized but unphosphorylated RB protein (Hensel et al., Cancer Res., 50: 3067-3072, 1990), which fails to form a complex with SV40 T antigen, suggesting that the RB gene of H209 may contain a subtle mutation. To define this mutation, the RB complementary DNA and genomic DNA were sequenced, revealing a point mutation in exon 21 that changed a G to a T. This results in an amino acid substitution of a Phe for Cys706. The mutant RB complementary DNA was used as a template for in vitro transcription and translation to synthesize the mutated protein. The resulting protein failed to bind to SV40 T antigen, demonstrating that a single missense mutation of the RB gene led to the complete inactivation of the ability of the RB protein to bind T antigen.
Bignon
Y J
YJ
Department of Pathology, School of Medicine, University of California, La Jolla 92093.
Shew
J Y
JY
Rappolee
D
D
Naylor
S L
SL
Lee
E Y
EY
Schnier
J
J
Lee
W H
WH
D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D001483:Base Sequence; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding / Q000235:genetics; D016160:Retinoblastoma Protein / Q000235:genetics* / Q000378:metabolism; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
D000952:Antigens, Polyomavirus Transforming; D016160:Retinoblastoma Protein